This is an application for continuation of a project whose major goal is to understand the regulation and function of the Ird3 kinase (IKK) complex and its subunits. Progress during the present funding period has been considerable - the different subunits of the IKK complex were molecularly identified and biochemically characterized, the genes encoding the different subunits were disrupted in mice and many of their physiological functions were identified. However, this progress has also generated important new questions that will be addressed during the next funding period. Most of the proposed work will focus on biochemical and functional characterization of the novel IKKc_-dependent NF-ld3 signaling pathway that functions through regulation of NF- _132/p100 processing, identified during the current period. We will study the function of this pathway in both B lymphocytes and in the splenic stroma. In addition to studying the biochemical basis for regulation of IKKa- dependent pl00 processing, we will identify the genes and the transcriptional factors that are targeted by this pathway and its role in autoimmune and lymphoproliferative diseases caused by overproduction of BAFF (B cell activating factor), a member of the TNF cytokine family. We will also continue to study the pathophysiological function of the canonical NF-Y,.B signaling pathway, whose activation depends on the IKK[3 subunit of the IKK complex. In this regard we will study the role of IKK_ in intestinal epithelial cells and macrophages in innate immunity, inflammation and cancer. We will also study the mechanisms (target genes) through which IKK_ exerts these functions. Finally, we will begin to study IKK-independent mechanisms for NF-rJ3 regulation. More specifically, we will study the role of the IKK-like NF-vJ3 activating kinase (NAK) in control of NF-rd3- dependent gone transcription, which may be exerted through phosphorylation of RelA and possibly other NF-Yd3 proteins. As before, our approach will incorporate biochemical studies, cell biology, the construction of genetically altered mouse strains and the use of specific disease models. In addition to basic information on the regulation of NF-v,_B signaling, these studies will generate much needed information regarding the pathophysiological functions of these pathways relevant to several major disease areas, including cancer, autoimmunity and inflammation.